Electroluminescent illumination source for optical detection systems

ABSTRACT

An optical detection system that utilizes an electroluminescent (EL) illumination source is provided. Unlike illumination sources used with some conventional optical detection systems, an EL device is relatively homogeneous and diffuse, and thus may provide uniform illumination to the test sample. In addition, the emitted light intensity of the EL device may be easily controlled by simply varying the voltage or the frequency of the applied current. The relatively flexibility of EL devices may also allow them to be readily incorporated into a chromatographic-based assay device for detecting the presence or absence of an analyte within a test sample.

RELATED APPLICATIONS

The present application claims priority to a provisional applicationhaving Ser. No. 60/608,941, which was filed on Mar. 30, 2004.

BACKGROUND OF THE INVENTION

Optical detection systems are often utilized to qualitatively,quantitatively, or semi-quantitatively determine the presence orconcentration of an analyte within a test sample. For example, someconventional optical detection systems employ reflective-based detectiontechniques in which the illumination source and detector are placed onthe same side of a test strip. Other conventional optical detectionsystems employ transmission-based detection techniques in which theillumination source and detector are placed on opposing sides of a teststrip. Transmission-based optical detection systems, for example,sometimes employ a photodiode detector positioned opposite to alight-emitting diode (LED) illumination source. Although such adetection system may provide certain benefits, it is often problematicin that the illumination source (e.g., LED) does not provide diffuselight. Thus, the system requires complex and expensive opticalcomponents, such as lenses or diffusers, to ensure that enough light issupplied to the test strip to provide an accurate result.

As such, a need currently exists for an optical detection system fordetecting the presence or quantity of an analyte that is inexpensive,easy to use, and accurate.

SUMMARY OF THE INVENTION

In accordance with one embodiment of the present invention, an opticaldetection system for detecting the presence or quantity of an analytewithin a test sample is disclosed. The system comprises anelectroluminescent illumination source that provides electromagneticradiation, which is capable of causing the production of a detectionsignal that correlates to the presence or quantity of the analyte.

In accordance with another embodiment of the present invention, anoptical detection system for detecting the presence or quantity of ananalyte within a test sample is disclosed. The system comprises an assaydevice that includes a porous membrane in communication with detectionprobes, the detection probes being capable of producing a detectionsignal. The system also comprises an electroluminescent illuminationsource capable of providing electromagnetic radiation that causes thedetection probes to produce the detection signal. A detector is alsoprovided that is capable of registering the detection signal produced bythe detection probes.

Other features and aspects of the present invention are discussed ingreater detail below.

BRIEF DESCRIPTION OF THE DRAWINGS

A full and enabling disclosure of the present invention, including thebest mode thereof, directed to one of ordinary skill in the art, is setforth more particularly in the remainder of the specification, whichmakes reference to the appended figure in which:

FIG. 1 is a cross-sectional view of one embodiment of anelectroluminescent (EL) device that may be used in the presentinvention; and

FIG. 2 is a schematic illustration of an optical detection system thatmay be employed in one embodiment of the present invention;

FIG. 3 schematically illustrates various embodiments of an opticaldetection system that may be used in the present invention, in whichFIG. 3 a illustrates an embodiment in which the illumination source anddetector are spaced relatively distant from the assay device; FIG. 3 billustrates the embodiment of FIG. 3 a in which an illumination lens anda detection lens are also used to focus light to and from the assaydevice; FIG. 3 c illustrates the embodiment of FIG. 3 b in which theillumination lens is removed and the illumination source is moved closerto the assay device; and FIG. 3 d illustrates the embodiment of FIG. 3 cin which the detection lens is removed and the detector is moved closerto the assay device;

FIG. 4 is a schematic illustration of another embodiment of an opticaldetection system of the present invention, which employs an ELillumination source;

FIG. 5 is a perspective view of one embodiment of a sample holder thatmay be used in the present invention, in which FIG. 5A shows the sampleholder prior to insertion of the assay strips; FIG. 5B shows the sampleholder in its open configuration with the strips inserted; and FIG. 5Cshows the sample holder in its closed configuration;

FIG. 6 is a perspective view of one embodiment of a cartridge in whichthe sample holder of FIG. 5 may be inserted;

FIG. 7 is a perspective view of one embodiment of an optical detectionsystem that utilizes the cartridge of FIG. 6 and the sample holder ofFIG. 5;

FIG. 8 is a perspective view of the optical detection system of FIG. 7contained within an enclosure; and

FIG. 9 graphically depicts the results of Example 1, in which the doseresponse is plotted versus CRP concentration (nanograms per milliliter).

Repeat use of reference characters in the present specification anddrawings is intended to represent same or analogous features or elementsof the invention.

DETAILED DESCRIPTION OF REPRESENTATIVE EMBODIMENTS Definitions

As used herein, the term “analyte” generally refers to a substance to bedetected. For instance, analytes may include antigenic substances,haptens, antibodies, and combinations thereof. Analytes include, but arenot limited to, toxins, organic compounds, proteins, peptides,microorganisms, amino acids, nucleic acids, hormones, steroids,vitamins, drugs (including those administered for therapeutic purposesas well as those administered for illicit purposes), drug intermediariesor byproducts, bacteria, virus particles and metabolites of orantibodies to any of the above substances. Specific examples of someanalytes include ferritin; creatinine kinase MB (CK-MB); digoxin;phenytoin; phenobarbitol; carbamazepine; vancomycin; gentamycin;theophylline; valproic acid; quinidine; luteinizing hormone (LH);follicle stimulating hormone (FSH); estradiol, progesterone; C-reactiveprotein; lipocalins; IgE antibodies; cytokines; vitamin B2micro-globulin; glycated hemoglobin (Gly. Hb); cortisol; digitoxin;N-acetylprocainamide NAPA); procainamide; antibodies to rubella, such asrubella-IgG and rubella IgM; antibodies to toxoplasmosis, such astoxoplasmosis IgG (Toxo-lgG) and toxoplasmosis IgM (Toxo-lgM);testosterone; salicylates; acetaminophen; hepatitis B virus surfaceantigen (HBsAg); antibodies to hepatitis B core antigen, such asanti-hepatitis B core antigen IgG and IgM (Anti-HBC); human immunedeficiency virus 1 and 2 (HIV 1 and 2); human T-cell leukemia virus 1and 2 (HTLV); hepatitis B e antigen (HBeAg); antibodies to hepatitis B eantigen (Anti-HBe); influenza virus; thyroid stimulating hormone (TSH);thyroxine (T4); total triiodothyronine (Total T3); free triiodothyronine(Free T3); carcinoembryoic antigen (CEA); lipoproteins, cholesterol, andtriglycerides; and alpha fetoprotein (AFP). Drugs of abuse andcontrolled substances include, but are not intended to be limited to,amphetamine; methamphetamine; barbiturates, such as amobarbital,secobarbital, pentobarbital, phenobarbital, and barbital;benzodiazepines, such as librium and valium; cannabinoids, such ashashish and marijuana; cocaine; fentanyl; LSD; methaqualone; opiates,such as heroin, morphine, codeine, hydromorphone, hydrocodone,methadone, oxycodone, oxymorphone and opium; phencyclidine; andpropoxyhene. Other potential analytes may be described in U.S. Pat. No.6,436,651 to Everhart, et al. and U.S. Pat. No. 4,366,241 to Tom et al.

As used herein, the term “test sample” generally refers to a biologicalmaterial suspected of containing the analyte. The test sample may bederived from any biological source, such as a physiological fluid,including, blood, interstitial fluid, saliva, ocular lens fluid,cerebral spinal fluid, sweat, urine, milk, ascites fluid, mucous, nasalfluid, sputum, synovial fluid, peritoneal fluid, vaginal fluid, menses,amniotic fluid, semen, and so forth. Besides physiological fluids, otherliquid samples may be used such as water, food products, and so forth,for the performance of environmental or food production assays. Inaddition, a solid material suspected of containing the analyte may beused as the test sample. The test sample may be used directly asobtained from the biological source or following a pretreatment tomodify the character of the sample. For example, such pretreatment mayinclude preparing plasma from blood, diluting viscous fluids, and soforth. Methods of pretreatment may also involve filtration,precipitation, dilution, distillation, mixing, concentration,inactivation of interfering components, the addition of reagents,lysing, etc. Moreover, it may also be beneficial to modify a solid testsample to form a liquid medium or to release the analyte.

Detailed Description

Reference now will be made in detail to various embodiments of theinvention, one or more examples of which are set forth below. Eachexample is provided by way of explanation of the invention, notlimitation of the invention. In fact, it will be apparent to thoseskilled in the art that various modifications and variations may be madein the present invention without departing from the scope or spirit ofthe invention. For instance, features illustrated or described as partof one embodiment, may be used on another embodiment to yield a stillfurther embodiment. Thus, it is intended that the present inventioncovers such modifications and variations as come within the scope of theappended claims and their equivalents.

In general, the present invention is directed to an optical detectionsystem that utilizes an electroluminescent (EL) illumination source.Unlike illumination sources used with some conventional opticaldetection systems, an EL device produces relatively homogeneous anddiffuse radiation, and thus may provide uniform illumination to the testsample. In addition, the intensity of radiation emitted by the EL devicemay be easily controlled by simply varying the voltage or the frequencyof the drive signal. Likewise, the wavelength of radiation may be easilyby controlled by varying the luminescent material. The relativelyflexibility of EL device may also allow it to be readily incorporatedinto a chromatographic-based assay device for detecting the presence orabsence of an analyte within a test sample.

I. Electroluminescent (EL) Device

An EL device is generally a capacitor structure that utilizes aluminescent material (e.g., phosphor particles) sandwiched betweenelectrodes, at least one of which is transparent to allow light toescape. Application of a voltage across the electrodes generates achanging electric field within the luminescent material that causes itto emit light. Generally speaking, any known EL device may be employedas the illumination source. For example, EL devices that employ“inorganic” or “organic” luminescent materials may be utilized in thepresent invention. Suitable “organic” EL devices include low and highmolecular weight devices. Likewise, suitable inorganic EL devicesinclude dispersion and thin-film phosphors. Dispersion EL devicesgenerally contain a dispersion of powder luminescent material in abinder, which is sandwiched between electrode layers. On the other hand,thin-film EL devices include a luminescent thin film that is sandwichedbetween a pair of insulating thin films and a pair of electrode layers,and is disposed on an electrically insulating substrate. Althoughcertainly not required, the dispersion-type EL devices are particularlydesired in certain embodiments of the present invention due to theirrelatively low cost and ease of manufacture.

Referring to FIG. 1, for instance, one embodiment of a dispersion-typeEL device 10 that may be used in the present invention is illustrated.As shown, the EL device 10 has a cathode 12, a dielectric layer 14, aluminescent layer 16, an anode 18, and a film 19. Additionalwater-impervious protective layers (not shown) may optionally be appliedto the cathode 12 and film 19 if desired. Leads 65 are electricallyattached to the respective cathode and anode layers 12 and 18. A drivercircuit (not shown) is connected to the leads via wiring, and the drivercircuit is connected to a power source (not shown). The details of thedriver circuit and power source generally depend on the requirements forthe particular EL device 10. For example, for relatively small ELdevices, e.g. having a 5-inch by 8-inch lit area, a low voltage circuitand battery power source may be used. Relatively large EL devices, onthe other hand, may be powered by a higher voltage circuit. The drivercircuit in an exemplary application converts DC voltage into an ACoutput for driving the EL device 10. Such AC inverters may generatearound 60 to 300 volts AC at 50 to 5000 Hertz. Driver circuits suitablefor this purpose are commercially available.

The cathode 12 may be formed from a metal (including metalloids) oralloys thereof (including intermetallic compounds). Examples of suitablematerials-for forming the cathode 12 include, but are not limited to,carbon; metals, such as aluminum, gold, silver, copper, platinum,palladium, iridium, and alloys thereof; and so forth. The thickness ofthe cathode 12 may generally vary, and may be deposited onto anelectrically insulating substrate (not shown). The substrate, forinstance, may be formed from ceramic materials, such as alumina (Al₂O₃),quartz glass (SiO₂), magnesia (MgO), forsterite (2MgO.SiO₂), steatite(MgO.SiO₂), mullite (3Al₂O₃.2SiO₂), beryllia (BeO), zirconia (ZrO₂),aluminum nitride (N), silicon nitride (SiN), silicon carbide (SiC),glass, heat resistant glass, and so forth. In addition, polymericmaterials may also be used to form the substrate, such as,polypropylene, polyethylene terephthalate, polyvinyl chloride,polymethylmethacrylate, and so forth.

The dielectric layer 14 is disposed on the cathode 12. The material ofwhich the dielectric layer 14 is formed may generally vary as is wellknown to those skilled in the art. For example, suitable materialsinclude, but are not limited to, perovskite structure dielectric andferroelectric materials, such as BaTiO₃, (Ba_(x)Ca_(1−x))TiO₃,(Ba_(x)Sr_(1−x))TiO₃, PbTiO₃ and Pb(Zr_(x)Ti_(1−x))O₃ (known as “PZT”);complex perovskite relaxation type ferroelectric materials, such asPb(Mg_(1/3)Nb_(2/3))O₃; bismuth layer compounds, such as Bi₄Ti₃O₁₂ andSrBi₂Ta₂O₉; and tungsten bronze type ferroelectric materials, such as(Sr_(x)Ba_(1−x))Nb₂O₆ and PbNb₂O₆. Still other suitable dielectricmaterials for use in the dielectric layer 14 may include dielectricmaterial, such as SiO₂, SiN, SiON, ZrO₂, Al₂O₃, Al₃N₄, Y₂O₃, Ta₂O₅, andso forth. In one particular embodiment, the dielectric layer 114 isformed from barium titanate (BaTiO₃).

The dielectric layer 14 may be formed using any of a variety oftechniques known to those skilled in the art. For example, thedielectric material used to form the layer 14 may first be admixed witha suitable solvent. Such solvents may include, for instance, glycolethers, alkyl ketones and aromatic solvents. Suitable glycol ethers mayinclude propylene glycol methyl ether, dipropylene glycol methyl ether,tripropylene glycol methyl ether, ethylene glycol ethyl ether,diethylene glycol butyl ether, and so forth. Suitable alkyl ketones mayinclude lower alkyl ketones, such as acetone, methyl ethyl ketone, ethylketone and methylisobutyl ketone, and so forth. Suitable aromaticsolvents may include toluene, xylene, and so forth. In one embodiment,barium titanate is added to a solvent in an amount from about 70% toabout 90% by weight. The barium titanate and the solvent are thenstirred together to form a homogeneous slurry.

Upon mixing with a solvent, the dielectric material may also be mixedwith a binder. For example, in some embodiments, the binder is added inan amount from about 10 to about 30 parts of the slurry. Suitablebinders are well known and include, for instance, epoxy resins,polystyrene, polyethylene, polyvinyl butyral, polyvinyl chloride,polyvinyl acetate, polyvinyl alcohol, polyesters, polyamides,polyacrylonitrile, polyacrylate, polymethylmethacrylate and the like. Insome embodiments, the binder is an adhesive thermoplastic reactionproduct of phenols and an excess of an epihalohydrin. Suitable phenolsinclude bisphenol A, dichlorobisphenol A, tetrachlorobisphenol A,tetrabromobisphenol A, bisphenol F and bisphenol ACP. The reaction iscarried out in the presence of a glycol ether or other suitable solvent.To this reaction product is added a resin such as a urethane or an epoxyresin in the range of from about 5 to 6 parts of resin to about 1 partof the epihalohydrin/phenol reaction product. Such binders are describedin more detail in U.S. Pat. No. 4,560,902 to Kardon and U.S. Pat. No.5,352,951 to Kardon. et al., which are incorporated herein in theirentirety by reference thereto for all purposes.

If desired, water may be added to the binder system at this step orfollowing assembly of the EL device 10. The water may be stirred intothe slurry before or after removal of the solvent. The amount of wateradded to the binder will vary somewhat in accordance with the amount ofwater the particular binder employed can absorb. For instance, at leastabout 1 part per million (“ppm”) (0.0001%) of water may be present andup to the maximum amount of water the binder will absorb. Cyanoethylpolyvinyl alcohol binders, for example, typically absorb a maximum ofabout 40,000 ppm (4.0%) of water. Cyanoalkylated pullulan binders, onthe other hand, typically absorb a maximum of about 100,000 ppm (10.0%)of water. However, in most cases, the amount of water added to thebinder is from about 500 ppm (0.05%) to about 20,000 ppm (2.0%). Thethickness of the resultant barium titanate/resin binder layer 114 istypically from about 0.2 to about 6 mils.

Referring again to FIG. 1, the EL device 10 also includes a luminescentlayer 16 disposed on the dielectric layer 14. The material of which theluminescent layer 16 may include phosphor particles. Suitable phosphorparticles may include a variety of metal oxide, sulfide, fluoride, andsilicate compounds. For example, such phosphor particles may includemanganese- and arsenic-activated zinc silicate (P39 phosphor),titanium-activated zinc silicate, manganese-activated zinc silicate (P1phosphor), cerium-activated yttrium silicate (P47 phosphor),manganese-activated magnesium silicate (P13 phosphor), lead- andmanganese-activated calcium silicate (P25 phosphor), terbium-activatedyttrium silicate, terbium-activated yttrium oxide, terbium-activatedyttrium aluminum oxide, terbium-activated gadolinium oxide,terbium-activated yttrium aluminum gallium oxide, europium-activatedyttrium oxide, europium-activated yttrium vanadium oxide,europium-activated yttrium oxysulfide, manganese-activated zinc sulfide,cesium-activated strontium sulfide, thulium-activated zinc sulfide,samarium-activated zinc sulfide, europium-activated calcium sulfide,terbium-activated zinc-sulfide, and cesium-activated calcium sulfide,and so forth.

The color emitted by the phosphor particles can be defined during themanufacture of the phosphor or by blending phosphors of different colorsto achieve composite color. Some specific examples of suitable phosphorsinclude manganese-activated zinc sulfide (yellowish orange lightemission), cesium-activated strontium sulfide (blue light emission),thulium-activated zinc sulfide (blue light emission), samarium-activatedzinc sulfide (red light emission), europium-activated calcium sulfide(red light emission), terbium-activated zinc-sulfide (green lightemission), and cesium-activated calcium sulfide (green light emission).

Phosphor particles typically have an average size of less than about 15micrometers, in some embodiments less than about 10 micrometers, and insome embodiments, less than about 5 micrometers. The luminescent layer16 may be formed using any of a variety of techniques known to thoseskilled in the art. For example, the encapsulated phosphor particles maybe admixed with a solvent, such as described above. The amount ofphosphor particles added to the solvent may range, for instance, fromabout 60% to about 95%, and in some embodiments, from about 75% to about85% by weight of the mixture. Likewise, after mixing, a binder, such asdescribed above, is also mixed with the phosphor particle slurry. Thebinder is typically present in an amount of from about 5 to about 40parts. If desired, the phosphor particles may also be encapsulatedwithin a protective material to form a water barrier as is well known inthe art. Suitable protective materials for encapsulating the phosphorparticles include, for instance, liquid crystals, polymeric binders,ceramic materials (e.g., colloidal silica, alumina, etc.), and so forth.Encapsulation techniques are described in more detail in U.S. Pat. No.4,097,776 to Allinikov; U.S. Pat. No. 4,513,023 to Wary; U.S. Pat. No.4,560,902 to Kardon; and U.S. Pat. No. 5,352,951 to Kardon, et al.,which are incorporated herein in their entirety by reference thereto forall purposes.

The phosphor particles preferably are deposited in a smooth, homogeneouslayer by any of a variety of techniques known to one of skill in theart. Such techniques include settling techniques, slurry methods (suchas screen printing, spin coating, and spin casting), electrophoresis, ordusting methods (such as electrostatic dusting, “phototacky” methods,and high pressure dusting). Settling techniques and slurry methodsinvolve forming a dispersion of the phosphor particles in a suitableliquid medium. One particularly desired deposition method is screenprinting. A suitable thickness for the phosphor/binder layer 16 whendried is about 0.2 to about 6 mils.

In addition to the layers mentioned above, the EL device 10 alsoincludes an anode 18 formed on a film 19, both of which are disposedover the luminescent layer 16. Desirably, the materials used for thelayers 18 and 19 are optically transparent. For example, the anode 18may be formed from a inorganic conductive oxide, such as indium oxide,indium tin oxide (ITO), tin oxide, and antimony tin oxide. In oneembodiment, an indium tin oxide (ITO) layer is utilized that has athickness of about 0.2 to 1 micrometers. Likewise, a suitable materialfor use as the film 19 may be a polymer film (e.g., polyester). Itshould be understood that the embodiments described above are merelyexemplary, and that any other known EL device may generally be used inthe present invention. For instance, other suitable EL devices aredescribed in U.S. Pat. No. 6,004,686 to Rasmussen, et al.; U.S. Pat. No.6,432,516 to Terasaki, et al.; U.S. Pat. No. 6,602,618 to Watanabe, etal.; U.S. Pat. No. 6,479,930 to Tanabe, et al.; U.S. Pat. No. 6,723,192to Nagano, et al.; and U.S. Pat. No. 6,734,469 to Yano, et al., as wellas U.S. patent application Publication Nos. 2003/0193289 to Shirakawa,et al.; 2004/0119400 to Takahashi. et al., and 2004/0070195 to Nelson,et al., all of which are incorporated herein in their entirety byreference thereto for all purposes.

II. Assay Device

In general, the assay device employed in the present invention mayperform any type of assay known in the art, including homogeneous andheterogeneous immunoassays. A homogeneous assay is an assay in whichuncomplexed labeled species are not separated from complexed labeledspecies. A heterogeneous assay is an assay in which uncomplexed labeledspecies are separated from complexed labeled species. Separation may becarried out by physical separation, e.g., by transferring one of thespecies to another reaction vessel, filtration, centrifugation,chromatography, solid phase capture, magnetic separation, and so forth,and may include one or more washing steps. The separation may also benonphysical in that no transfer of one or both of the species isconducted, but the species are separated from one another in situ. Inone particular embodiment, for example, a heterogeneous immunoassay isutilized. Such immunoassays utilize mechanisms of the immune systems,wherein antibodies are produced in response to the presence of antigensthat are pathogenic or foreign to the organisms. These antibodies andantigens, i.e., immunoreactants, are capable of binding with oneanother, thereby causing a highly specific reaction mechanism that maybe used to determine the presence or concentration of that particularantigen in a fluid test sample.

Referring to FIG. 2, for example, a chromatographic-based assay device20 that is configured to perform a heterogeneous immunoassay will now bedescribed in more detail. As shown, the assay device 20 contains achromatographic medium 23 having a first surface 13 and an opposingsecond surface 15. The first surface 13 of the medium 23 is positionedadjacent to a support 21. The chromatographic medium 23 is generallymade from a material through which the test sample is capable ofpassing, such as a fluidic channel, porous membrane, etc. Likewise, themedium 23 is also made from a material through which electromagneticradiation may transmit, such as an optically diffuse (e.g.,transluscent) or transparent material. In one particular embodiment, forexample, the chromatographic medium 23 is made from an optically diffuseporous membrane formed from materials such as, but not limited to,natural, synthetic, or naturally occurring materials that aresynthetically modified, such as polysaccharides (e.g., cellulosematerials such as paper and cellulose derivatives, such as celluloseacetate and nitrocellulose); polyether sulfone; polyethylene; nylon;polyvinylidene fluoride (PVDF); polyester; polypropylene; silica;inorganic materials, such as deactivated alumina, diatomaceous earth,MgSO₄, or other inorganic finely divided material uniformly dispersed ina porous polymer matrix, with polymers such as vinyl chloride, vinylchloride-propylene copolymer, and vinyl chloride-vinyl acetatecopolymer; cloth, both naturally occurring (e.g., cotton) and synthetic(e.g., nylon or rayon); porous gels, such as silica gel, agarose,dextran, and gelatin; polymeric films, such as polyacrylamide; and soforth. In one particular embodiment, the chromatographic medium 23 isformed from nitrocellulose and/or polyether sulfone materials. It shouldbe understood that the term “nitrocellulose” refers to nitric acidesters of cellulose, which may be nitrocellulose alone, or a mixed esterof nitric acid and other acids, such as aliphatic carboxylic acidshaving from 1 to 7 carbon atoms.

The size and shape of the chromatographic medium 23 may generally varyas is readily recognized by those skilled in the art. For instance, aporous membrane strip may have a length of from about 10 to about 100millimeters, in some embodiments from about 20 to about 80 millimeters,and in some embodiments, from about 40 to about 60 millimeters. Thewidth of the membrane strip may also range from about 0.5 to about 20millimeters, in some embodiments from about 1 to about 15 millimeters,and in some embodiments, from about 2 to about 10 millimeters. Likewise,the thickness of the membrane strip is generally small enough to allowtransmission-based detection. For example, the membrane strip may have athickness less than about 500 micrometers, in some embodiments less thanabout 250 micrometers, and in some embodiments, less than about 150micrometers.

As stated above, the support 21 carries the chromatographic medium 23.For example, the support 21 may be positioned directly adjacent to thechromatographic medium 23 as shown in FIG. 2, or one or more interveninglayers may be positioned between the chromatographic medium 23 and thesupport 21. Regardless, the support 21 may generally be formed from anymaterial able to carry the chromatographic medium 23. Generally, thesupport 21 is formed from a material that is transmissive to light, suchas transparent or optically diffuse (e.g., transluscent) materials.Also, it is generally desired that the support 21 is liquid-impermeableso that fluid flowing through the medium 23 does not leak through thesupport 21. Examples of suitable materials for the support include, butare not limited to, glass; polymeric materials, such as polystyrene,polypropylene, polyester (e.g., Mylar® film), polybutadiene,polyvinylchloride, polyamide, polycarbonate, epoxides, methacrylates,and polymelamine; and so forth. To provide a sufficient structuralbacking for the chromatographic medium 23, the support 21 is generallyselected to have a certain minimum thickness. Likewise, the thickness ofthe support 21 is typically not so large as to adversely affect itsoptical properties. Thus, for example, the support 21 may have athickness that ranges from about 100 to about 5,000 micrometers, in someembodiments from about 150 to about 2,000 micrometers, and in someembodiments, from about 250 to about 1,000 micrometers. For instance,one suitable membrane strip having a thickness of about 125 micrometersmay be obtained from Millipore Corp. of Bedford, Mass. under the name“SHF180UB25.”

As is well known the art, the chromatographic medium 23 may be cast ontothe support 21, wherein the resulting laminate may be die-cut to thedesired size and shape. Alternatively, the chromatographic medium 23 maysimply be laminated to the support 21 with, for example, an adhesive. Insome embodiments, a nitrocellulose or nylon porous membrane is adheredto a Mylar® film. An adhesive is used to bind the porous membrane to theMylar® film, such as a pressure-sensitive adhesive. Laminate structuresof this type are believed to be commercially available from MilliporeCorp. of Bedford, Mass. Still other examples of suitable laminate assaydevice structures are described in U.S. Pat. No. 5,075,077 to Durley,III, et al., which is incorporated herein in its entirety by referencethereto for all purposes.

The selection of an adhesive for laminating the support 21, thechromatographic medium 23, and/or any other layer of the device maydepend on a variety of factors, including the desired optical propertiesof the detection system and the materials used to form the assay device.For example, in some embodiments, the selected adhesive is opticallytransparent and compatible with the chromatographic medium 23 andsupport 21. Optical transparency may minimize any adverse affect thatthe adhesive might otherwise have on the optical detection system.Suitable optically transparent adhesives may be formed, for instance,from acrylate or (meth)acrylate polymers, such as polymers of(meth)acrylate esters, acrylic or (meth)acrylic acid monomers, and soforth. Exemplary (meth)acrylate ester monomers include monofunctionalacrylate or methacrylate esters of non-tertiary alkyl alcohols, such asmethyl acrylate, ethyl acrylate, propyl acrylate, n-butyl acrylate,isobutyl acrylate, 2-methylbutyl acrylate, 2-ethylhexyl acrylate,2-ethylhexyl methacrylate, n-octyl acrylate, n-octyl methacrylate,isooctyl acrylate, isooctyl methacrylate, isononyl acrylate, isodecylacrylate, isobomyl acrylate, isobornyl methacrylate, vinyl acetate, andmixtures thereof. Exemplary (meth)acrylic acid monomers include acrylicacid, methacrylic acid, beta-carboxyethyl acrylate, itaconic acid,crotonic acid, fumaric acid, and so forth. Several examples of suchoptically transparent adhesives are described in U.S. Pat. No. 6,759,121to Alahapperuma, et al., which is incorporated herein in its entirety byreference thereto for all purposes. Further, suitable transparentadhesives may also be obtained from Adhesives Research, Inc. of GlenRock, Pa. under the name ARclear® 8154, which is an unsupportedoptically clear acrylic pressure-sensitive adhesive. Other suitabletransparent adhesives may be obtained from 3M Corp. of St. Paul, Minn.under the names “9843” or “8146.” In addition, the manner in which theadhesive is applied may also enhance the optical properties of the assaydevice. For instance, the adhesive may enhance certain opticalproperties of the support (e.g., diffusiveness). Thus, in one particularembodiment, such an adhesive may be applied in a pattern thatcorresponds to the areas in which enhanced optical properties aredesired.

Referring again to FIG. 2, an absorbent pad 28 is provided on the secondsurface 15 that generally receives fluid after it migrates through theentire chromatographic medium 23 As is well known in the art, theabsorbent pad 28 may also assist in promoting capillary action and fluidflow through the chromatographic medium 23 To initiate the detection ofan analyte within the test sample, a user may directly apply the testsample to a portion of the chromatographic medium 23 through which itmay then travel in the direction illustrated by arrow “L” in FIG. 2.Alternatively, the test sample may first be applied to a sample pad (notshown) that is in fluid communication with the chromatographic medium23. Some suitable materials that may be used to form the absorbent pad28 and/or sample pad include, but are not limited to, nitrocellulose,cellulose, porous polyethylene pads, and glass fiber filter paper. Ifdesired, the sample pad may also contain one or more assay pretreatmentreagents, either diffusively or non-diffusively attached thereto.

In the illustrated embodiment, the test sample travels from the samplepad (not shown) to a conjugate pad 22 that is placed in communicationwith one end of the sample pad. The conjugate pad 22 is formed from amaterial through which a fluid is capable of passing. For example, inone embodiment, the conjugate pad 22 is formed from glass fibers.Although only one conjugate pad 22 is shown, it should be understoodthat other conjugate pads may also be used in the present invention.

To facilitate accurate detection of the presence or absence of ananalyte within the test sample, a predetermined amount of detectionprobes may be applied at various locations of the assay device 20. Suchdetection probes contain a substance that directly or indirectlyproduces an optically detectable signal, such as molecules, polymers,dendrimers, and so forth. Suitable detectable substances may include,for instance, luminescent compounds (e.g., fluorescent, phosphorescent,etc.); radioactive compounds; visual compounds (e.g., colored dye ormetallic substance, e.g., gold); liposomes or other vesicles containingsignal-producing substances; enzymes and/or substrates, and so forth.Other suitable detectable substances may be described in U.S. Pat. No.5,670,381 to Jou,. et al. and U.S. Pat. No. 5,252,459 to Tarcha, et al.,which are incorporated herein in their entirety by reference thereto forall purposes. If the detectable substance is colored, the idealelectromagnetic radiation is light of a complementary wavelength. Forinstance, blue detection probes strongly absorb red light.

In some embodiments, the detectable substance may be a luminescentcompound that produces an optically detectable signal. For example,suitable fluorescent molecules may include, but not limited to,fluorescein, europium chelates, phycobiliprotein, rhodamine, and theirderivatives and analogs. Other suitable fluorescent compounds aresemiconductor nanocrystals commonly referred to as “quantum dots.” Forexample, such nanocrystals may contain a core of the formula CdX,wherein X is Se, Te, S, and so forth. The nanocrystals may also bepassivated with an overlying shell of the formula YZ, wherein Y is Cd orZn, and Z is S or Se. Other examples of suitable semiconductornanocrystals may also be described in U.S. Pat. No. 6,261,779 toBarbera-Guillem, et al. and U.S. Pat. No. 6,585,939 to Dapprich, whichare incorporated herein in their entirety by reference thereto for allpurposes.

Further, suitable phosphorescent compounds may include metal complexesof one or more metals, such as ruthenium, osmium, rhenium, iridium,rhodium, platinum, indium, palladium, molybdenum, technetium, copper,iron, chromium, tungsten, zinc, and so forth. Especially preferred areruthenium, rhenium, osmium, platinum, and palladium. The metal complexmay contain one or more ligands that facilitate the solubility of thecomplex in an aqueous or nonaqueous environment. For example, somesuitable examples of ligands include, but are not limited to, pyridine;pyrazine; isonicotinamide; imidazole; bipyridine; terpyridine;phenanthroline; dipyridophenazine; porphyrin, porphine, and derivativesthereof. Such ligands may be, for instance, substituted with alkyl,substituted alkyl, aryl, substituted aryl, aralkyl, substituted aralkyl,carboxylate, carboxaldehyde, carboxamide, cyano, amino, hydroxy, imino,hydroxycarbonyl, aminocarbonyl, amidine, guanidinium, ureide,sulfur-containing groups, phosphorus containing groups, and thecarboxylate ester of N-hydroxy-succinimide.

Porphyrins and porphine metal complexes possess pyrrole groups coupledtogether with methylene bridges to form cyclic structures with metalchelating inner cavities. Many of these molecules exhibit strongphosphorescence properties at room temperature in suitable solvents(e.g., water) and an oxygen-free environment. Some suitable porphyrincomplexes that are capable of exhibiting phosphorescent propertiesinclude, but are not limited to, platinum (II) coproporphyrin-I and III,palladium (II) coproporphyrin, ruthenium coproporphyrin,zinc(II)-coproporphyrin-I, derivatives thereof, and so forth. Similarly,some suitable porphine complexes that are capable of exhibitingphosphorescent properties include, but not limited to, platinum(II)tetra-meso-fluorophenylporphine and palladium(II)tetra-meso-fluorophenylporphine. Still other suitable porphyrin and/orporphine complexes are described in U.S. Pat. No. 4,614,723 to Schmidt,et al.; U.S. Pat. No. 5,464,741 to Hendrix; U.S. Pat. No. 5,518,883 toU.S. Pat. No. Soini; 5,922,537 to Ewart, et al.; U.S. Pat. No. 6,004,530to Sagner, et al.; and U.S. Pat. No. 6,582,930 to Ponomarev, et al.,which are incorporated herein in their entirety by reference thereto forall purposes.

Bipyridine metal complexes may also be utilized as phosphorescentcompounds. Some examples of suitable bipyridine complexes include, butare note limited to, bis[(4,4′-carbomethoxy)-2,2′-bipyridine]2-[3-(4-methyl-2,2′-bipyridine4-yl)propyl]-1,3-dioxolane ruthenium (II);bis(2,2′bipyridine)[4-(butan-1-al)-4′-methyl-2,2′-bi-pyridine]ruthenium(II); bis(2,2′-bipyridine)[4-(4′-methyl-2,2′-bipyridine-4′-yl)-butyricacid] ruthenium (II); tris(2,2′bipyridine)ruthenium (II);(2,2′-bipyridine) [bis-bis(1,2-diphenylphosphino)ethylene]2-[3-(4-methyl-2,2′-bipyridine-4′-yl)propyl]-1,3-dioxolane osmium (II);bis(2,2′-bipyridine)[4-(4′-methyl-2,2′-bipyridine)-butylamine]ruthenium(II);bis(2,2′-bipyridine)[1-bromo4(4′-methyl-2,2′-bipyridine4-yl)butane]ruthenium(II); bis(2,2′-bipyridine)maleimidohexanoic acid,4-methyl-2,2′-bipyridine-4′-butylamide ruthenium (II), and so forth.Still other suitable metal complexes that may exhibit phosphorescentproperties may be described in U.S. Pat. No. 6,613,583 to Richter, etal.; U.S. Pat. No. 6,468,741 to Massey, et al.; U.S. Pat. No. 6,444,423to Meade, et al.; U.S. Pat. No. 6,362,011 to Massey, et al.; U.S. Pat.No. 5,731,147 to Bard, et al.; and U.S. Pat. No. 5,591,581 to Massey, etal., which are incorporated herein in their entirety by referencethereto for all purposes.

In some cases, luminescent compounds may have a relatively long emissionlifetime may have a relatively large “Stokes shift.” The term “Stokesshift” is generally defined as the displacement of spectral lines orbands of luminescent radiation to a longer emission wavelength than theexcitation lines or bands. A relatively large Stokes shift allows theexcitation wavelength of a luminescent compound to remain far apart fromits emission wavelengths and is desirable because a large differencebetween excitation and emission wavelengths makes it easier to eliminatethe reflected excitation radiation from the emitted signal. Further, alarge Stokes shift also minimizes interference from luminescentmolecules in the sample and/or light scattering due to proteins orcolloids, which are present with some body fluids (e.g., blood). Inaddition, a large Stokes shift also minimizes the requirement forexpensive, high-precision filters to eliminate background interference.For example, in some embodiments, the luminescent compounds have aStokes shift of greater than about 50 nanometers, in some embodimentsgreater than about 100 nanometers, and in some embodiments, from about100 to about 350 nanometers.

For example, exemplary fluorescent compounds having a large Stokes shiftinclude lanthanide chelates of samarium (Sm (III)), dysprosium (Dy(III)), europium (Eu (III)), and terbium (Tb (III)). Such chelates mayexhibit strongly red-shifted, narrow-band, long-lived emission afterexcitation of the chelate at substantially shorter wavelengths.Typically, the chelate possesses a strong ultraviolet excitation banddue to a chromophore located close to the lanthanide in the molecule.Subsequent to excitation by the chromophore, the excitation energy maybe transferred from the excited chromophore to the lanthanide. This isfollowed by a fluorescence emission characteristic of the lanthanide.Europium chelates, for instance, have Stokes shifts of about 250 toabout 350 nanometers, as compared to only about 28 nanometers forfluorescein. Also, the fluorescence of europium chelates is long-lived,with lifetimes of about 100 to about 1000 microseconds, as compared toabout 1 to about 100 nanoseconds for other fluorescent labels. Inaddition, these chelates have a narrow emission spectra, typicallyhaving bandwidths less than about 10 nanometers at about 50% emission.One suitable europium chelate is N-(p-isothiocyanatobenzyl)-diethylenetriamine tetraacetic acid-Eu⁺³.

In addition, lanthanide chelates that are inert, stable, andintrinsically fluorescent in aqueous solutions or suspensions may alsobe used in the present invention to negate the need for micelle-formingreagents, which are often used to protect chelates having limitedsolubility and quenching problems in aqueous solutions or suspensions.One example of such a chelate is 4-[2-(4-isothiocyanatophenyl)ethynyl]-2,6-bis([N, N-bis(carboxymethyl )amino]methyl)-pyridine [Ref:Lovgren, T., et al.; Clin. Chem. 42,1196-1201 (1996)]. Severallanthanide chelates also show exceptionally high signal-to-noise ratios.For example, one such chelate is a tetradentate β-diketonate-europiumchelate [Ref: Yuan, J. and Matsumoto, K.; Anal. Chem. 70, 596-601(1998)]. In addition to the fluorescent labels described above, otherlabels that are suitable for use in the present invention may bedescribed in U.S. Pat. No. 6,030,840 to Mullinax, et al.; U.S. Pat. No.5,585,279 to Davidson; U.S. Pat. No. 5,573,909 to Singer, et al.; U.S.Pat. No. 6,242,268 to Wieder, et al.; and U.S. Pat. No. 5,637,509 toHemmila, et al., which are incorporated herein in their entirety byreference thereto for all purposes.

Detectable substances, such as described above, may be used alone or inconjunction with a particle (sometimes referred to as “beads” or“microbeads”). For instance, naturally occurring particles, such asnuclei, mycoplasma, plasmids, plastids, mammalian cells (e.g.,erythrocyte ghosts), unicellular microorganisms (e.g., bacteria),polysaccharides (e.g., agarose), etc., may be used. Further, syntheticparticles may also be utilized. For example, in one embodiment, latexmicroparticles that are labeled with a fluorescent or colored dye areutilized. Although any synthetic particle may be used in the presentinvention, the particles are typically formed from polystyrene,butadiene styrenes, styreneacrylic-vinyl terpolymer,polymethylmethacrylate, polyethylmethacrylate, styrene-maleic anhydridecopolymer, polyvinyl acetate, polyvinylpyridine, polydivinylbenzene,polybutyleneterephthalate, acrylonitrile, vinylchloride-acrylates, andso forth, or an aldehyde, carboxyl, amino, hydroxyl, or hydrazidederivative thereof. Other suitable particles may be described in U.S.Pat. No. 5,670,381 to Jou, et al. and U.S. Pat. No. 5,252,459 to Tarcha,et al. Commercially available examples of suitable fluorescent particlesinclude fluorescent carboxylated microspheres sold by Molecular Probes,Inc. under the trade names “FluoSphere” (Red 580/605) and“TransfluoSphere” (543/620), as well as “Texas Red” and 5- and6-carboxytetramethylrhodamine, which are also sold by Molecular Probes,Inc. In addition, commercially available examples of suitable colored,latex microparticles include carboxylated latex beads sold by Bang'sLaboratory, Inc. Metallic particles (e.g., gold particles) may also beutilized in the present invention.

When utilized, the shape of the particles may generally vary. In oneparticular embodiment, for instance, the particles are spherical inshape. However, it should be understood that other shapes are alsocontemplated by the present invention, such as plates, rods, discs,bars, tubes, irregular shapes, etc. In addition, the size of theparticles may also vary. For instance, the average size (e.g., diameter)of the particles may range from about 0.1 nanometers to about 1,000microns, in some embodiments, from about 0.1 nanometers to about 100microns, and in some embodiments, from about 1 nanometer to about 10microns. For instance, “micron-scale” particles are often desired. Whenutilized, such “micron-scale” particles may have an average size of fromabout 1 micron to about 1,000 microns, in some embodiments from about 1micron to about 100 microns, and in some embodiments, from about 1micron to about 10 microns. Likewise, “nano-scale” particles may also beutilized. Such “nano-scale” particles may have an average size of fromabout 0.1 to about 10 nanometers, in some embodiments from about 0.1 toabout 5 nanometers, and in some embodiments, from about 1 to about 5nanometers.

In some instances, it may be desired to modify the detection probes insome manner so that they are more readily able to bind to the analyte.In such instances, the detection probes may be modified with certainspecific binding members that are adhered thereto to form conjugatedprobes. Specific binding members generally refer to a member of aspecific binding pair, i.e., two different molecules where one of themolecules chemically and/or physically binds to the second molecule. Forinstance, immunoreactive specific binding members may include antigens,haptens, aptamers, antibodies (primary or secondary), and complexesthereof, including those formed by recombinant DNA methods or peptidesynthesis. An antibody may be a monoclonal or polyclonal antibody, arecombinant protein or a mixture(s) or fragment(s) thereof, as well as amixture of an antibody and other specific binding members. The detailsof the preparation of such antibodies and their suitability for use asspecific binding members are well known to those skilled in the art.Other common specific binding pairs include but are not limited to,biotin and avidin (or derivatives thereof), biotin and streptavidin,carbohydrates and lectins, complementary nucleotide sequences (includingprobe and capture nucleic acid sequences used in DNA hybridizationassays to detect a target nucleic acid sequence), complementary peptidesequences including those formed by recombinant methods, effector andreceptor molecules, hormone and hormone binding protein, enzymecofactors and enzymes, enzyme inhibitors and enzymes, and so forth.Furthermore, specific binding pairs may include members that are analogsof the original specific binding member. For example, a derivative orfragment of the analyte, i.e., an analyte-analog, may be used so long asit has at least one epitope in common with the analyte.

The specific binding members may generally be attached to the detectionprobes using any of a variety of well-known techniques. For instance,covalent attachment of the specific binding members to the detectionprobes (e.g., particles) may be accomplished using carboxylic, amino,aldehyde, bromoacetyl, iodoacetyl, thiol, epoxy and other reactive orlinking functional groups, as well as residual free radicals and radicalcations, through which a protein coupling reaction may be accomplished.A surface functional group may also be incorporated as a functionalizedco-monomer because the surface of the detection probe may contain arelatively high surface concentration of polar groups. In addition,although detection probes are often functionalized after synthesis, suchas with poly(thiophenol), the detection probes may be capable of directcovalent linking with a protein without the need for furthermodification. For example, in one embodiment, the first step ofconjugation is activation of carboxylic groups on the probe surfaceusing carbodiimide. In the second step, the activated carboxylic acidgroups are reacted with an amino group of an antibody to form an amidebond. The activation and/or antibody coupling may occur in a buffer,such as phosphate-buffered saline (PBS) (e.g., pH of 7.2) or2-(N-morpholino) ethane sulfonic acid (MES) (e.g., pH of 5.3). Theresulting detection probes may then be contacted with ethanolamine, forinstance, to block any remaining activated sites. Overall, this processforms a conjugated detection probe, where the antibody is covalentlyattached to the probe. Besides covalent bonding, other attachmenttechniques, such as physical adsorption, may also be utilized in thepresent invention.

Referring again to FIG. 2, the chromatographic medium 23 also defines adetection zone 31 within which is immobilized a receptive material thatis capable of binding to the conjugated detection probes. For example,in some embodiments, the receptive material may be a biologicalreceptive material. Such biological receptive materials are well knownin the art and may include, but are not limited to, antigens, haptens,protein A or G, neutravidin, avidin, streptavidin, captavidin, primaryor secondary antibodies (e.g., polyclonal, monoclonal, etc.), andcomplexes thereof. In many cases, it is desired that these biologicalreceptive materials are capable of binding to a specific binding member(e.g., antibody) present on the detection probes. The receptive materialserves as a stationary binding site for complexes formed between theanalyte and conjugated detection probes. Specifically, analytes, such asantibodies, antigens, etc., typically have two or more binding sites(e.g., epitopes). Upon reaching the detection zone 31, one of thesebinding sites is occupied by the specific binding member of theconjugated probe. However, the free binding site of the analyte may bindto the immobilized receptive material. Upon being bound to theimmobilized receptive material, the complexed probes form a new ternarysandwich complex.

The detection zone 31 may generally provide any number of distinctdetection regions so that a user may better determine the concentrationof a particular analyte within a test sample. Each region may containthe same receptive materials, or may contain different receptivematerials for capturing multiple analytes. For example, the detectionzone 31 may include two or more distinct detection regions (e.g., lines,dots, etc.). The detection regions may be disposed in the form of linesin a direction that is substantially perpendicular to the flow of thetest sample through the assay device 20. Likewise, in some embodiments,the detection regions may be disposed in the form of lines in adirection that is substantially parallel to the flow of the test samplethrough the assay device 20.

Although the detection zone 31 provides accurate results for detectingan analyte, it is sometimes difficult to determine the relativeconcentration of the analyte within the test sample under actual testconditions. Thus, the assay device 20 may also include a calibrationzone 32. In this embodiment, the calibration zone 32 is positioneddownstream from the detection zone 31. Alternatively, however, thecalibration zone 32 may also be positioned upstream from the detectionzone 31. The calibration zone 32 may be provided with a receptivematerial that is capable of binding to calibration probes or uncomplexeddetection probes that pass through the length of the chromatographicmedium 23 When utilized, the calibration probes may be formed from thesame or different materials as the detection probes. Generally speaking,the calibration probes are selected in such a manner that they do notbind to the receptive material at the detection zone 31.

The receptive material of the calibration zone 32 may be the same ordifferent than the receptive material used in the detection zone 31. Forexample, in one embodiment, the receptive material is a biologicalreceptive material. In addition, it may also be desired to utilizevarious non-biological materials for the receptive material of thecalibration zone 32. The polyelectrolytes may have a net positive ornegative charge, as well as a net charge that is generally neutral. Forinstance, some suitable examples of polyelectrolytes having a netpositive charge include, but are not limited to, polylysine(commercially available from Sigma-Aldrich Chemical Co., Inc. of St.Louis, Mo.), polyethylenimine; epichlorohydrin-functionalized polyaminesand/or polyamidoamines, such as poly(dimethylamine-co-epichlorohydrin);polydiallyldimethyl-ammonium chloride; cationic cellulose derivatives,such as cellulose copolymers or cellulose derivatives grafted with aquaternary ammonium water-soluble monomer; and so forth. In oneparticular embodiment, CelQuat® SC-230M or H-100 (available fromNational Starch & Chemical, Inc.), which are cellulosic derivativescontaining a quaternary ammonium water-soluble monomer, may be utilized.Moreover, some suitable examples of polyelectrolytes having a netnegative charge include, but are not limited to, polyacrylic acids, suchas poly(ethylene-co-methacrylic acid, sodium salt), and so forth. Itshould also be understood that other polyelectrolytes may also beutilized in the present invention, such as amphiphilic polyelectrolytes(i.e., having polar and non-polar portions). For instance, some examplesof suitable amphiphilic polyelectrolytes include, but are not limitedto, poly(styryl-b-N-methyl 2-vinyl pyridinium iodide) andpoly(styryl-b-acrylic acid), both of which are available from PolymerSource, Inc. of Dorval, Canada. Further examples of internal calibrationsystems that utilize polyelectrolytes are described in more detail inU.S. Patent app. Publication No. 2003/0124739 to Song, et al., which isincorporated herein in it entirety by reference thereto for allpurposes.

In some cases, the chromatographic medium 23 may also define a controlzone (not shown) that gives a signal to the user that the assay isperforming properly. For instance, the control zone (not shown) maycontain an immobilized receptive material that is generally capable offorming a chemical and/or physical bond with probes or with thereceptive material immobilized on the probes. Some examples of suchreceptive materials include, but are not limited to, antigens, haptens,antibodies, protein A or G, avidin, streptavidin, secondary antibodies,and complexes thereof. In addition, it may also be desired to utilizevarious non-biological materials for the control zone receptivematerial. For instance, in some embodiments, the control zone receptivematerial may also include a polyelectrolyte, such as described above,that may bind to uncaptured probes. Because the receptive material atthe control zone is only specific for probes, a signal forms regardlessof whether the analyte is present. The control zone may be positioned atany location along the medium 23, but is typically positioned upstreamfrom the detection zone 31.

Various formats may be used to test for the presence or absence of ananalyte using the assay device 20. For instance, a “sandwich” formattypically involves mixing the test sample with detection probesconjugated with a specific binding member (e.g., antibody) for theanalyte to form complexes between the analyte and the conjugated probes.These complexes are then allowed to contact a receptive material (e.g.,antibodies) immobilized within the detection zone. Binding occursbetween the analyte/probe conjugate complexes and the immobilizedreceptive material, thereby localizing “sandwich” complexes that aredetectable to indicate the presence of the analyte. This technique maybe used to obtain quantitative or semi-quantitative results. Someexamples of such sandwich-type assays are described by U.S. Pat. No.4,168,146 to Grubb, et al. and U.S. Pat. No. 4,366,241 to Tom, et al.,which are incorporated herein in their entirety by reference thereto forall purposes. In a competitive assay, the labeled probe is generallyconjugated with a molecule that is identical to, or an analog of, theanalyte. Thus, the labeled probe competes with the analyte of interestfor the available receptive material. Competitive assays are typicallyused for detection of analytes such as haptens, each hapten beingmonovalent and capable of binding only one antibody molecule. Examplesof competitive immunoassay devices are described in U.S. Pat. No.4,235,601 to Deutsch, et al., U.S. Pat. No. 4,442,204 to Liotta, andU.S. Pat. No. 5,208,535 to Buechler, et al., which are incorporatedherein in their entirety by reference thereto for all purposes. Variousother device configurations and/or assay formats are also described inU.S. Pat. No. 5,395,754 to Lambotte, et al.; U.S. Pat. No. 5,670,381 toJou, et al.; and U.S. Pat. No. 6,194,220 to Malick, et al., which areincorporated herein in their entirety by reference thereto for allpurposes.

III. Optical Detection Systems

When utilized as an illumination source of an optical detection system,EL devices may provide a variety of benefits. For instance, unlike pointlight sources used with many conventional optical detection systems(e.g. LEDs), EL devices emit relatively homogeneous and diffuse light,and may thus provide uniform illumination. This may eliminate the needfor additional diffusers often required in point-source illuminationsystems. In addition, the light intensity emitted by EL device may beeasily controlled by simply varying the voltage or the frequency of thedrive signal. Thus, an EL device allows for the use of optical readersthat are relatively simple, portable, and inexpensive.

The actual configuration and structure of the optical detection systemin which the EL device is employed may generally vary as is readilyunderstood by those skilled in the art. For example, optical detectiontechniques that may be utilized include, but are not limited to,luminescence (e.g., fluorescence, phosphorescence, etc.), absorbance(e.g., fluorescent or non-fluorescent), diffraction, etc. Typically, theoptical system is capable of emitting light and also registering adetection signal (e.g., transmitted or reflected light, emittedfluorescence or phosphorescence, etc.). For example, in one embodiment,a reflectance spectrophotometer may be utilized to detect the presenceof probes that exhibit a visual color (e.g. dyed latex microparticles).One suitable reflectance spectrophotometer is described, for instance,in U.S. patent app. Pub. No. 2003/0119202 to Kaylor, et al., which isincorporated herein in its entirety by reference thereto for allpurposes. In another embodiment, a reflectance-mode spectrofluorometermay be used to detect the presence of probes that exhibit fluorescence.Suitable spectrofluororheters and related detection techniques aredescribed, for instance, in U.S. patent app. Pub. No. 2004/0043502 toSonq, et al., which is incorporated herein in its entirety by referencethereto for all purposes. Likewise, a transmission-mode detection systemmay also be used to detect the presence of detection probes.

Referring again to FIG. 2, for example, one embodiment of atransmission-based optical detection system is shown that employs anelectroluminescent (EL) illumination source 52 and a detector 54 Thedetector 54 may generally be any device known in the art that is capableof sensing an optical signal. For instance, the detector 54 may be anelectronic imaging detector that is configured for spatialdiscrimination. Some examples of such electronic imaging sensors includehigh speed, linear charge-coupled devices (CCD), charge-injectiondevices (CID), complementary-metal-oxide-semiconductor (CMOS) devices,and so forth. Such image detectors, for instance, are generallytwo-dimensional arrays of electronic light sensors, although linearimaging detectors (e.g., linear CCD detectors) that include a singleline of detector pixels or light sensors, such as, for example, thoseused for scanning images, may also be used. Each array includes a set ofknown, unique positions that may be referred to as “addresses.” Eachaddress in an image detector is occupied by a sensor that covers an area(e.g., an area typically shaped as a box or a rectangle). This area isgenerally referred to as a “pixel” or pixel area. A detector pixel, forinstance, may be a CCD, CID, or a CMOS sensor, or any other device orsensor that detects or measures light. The size of detector pixels mayvary widely, and may in some cases have a diameter or length as low as0.2 micrometers.

In other embodiments, the detector 54 may be a light sensor that lacksspatial discrimination capabilities. For instance, examples of suchlight sensors may include photomultiplier devices, photodiodes, such asavalanche photodiodes or silicon photodiodes, and so forth. Siliconphotodiodes are sometimes advantageous in that they are inexpensive,sensitive, capable of high-speed operation (short risetime/highbandwidth), and easily integrated into most other semiconductortechnology and monolithic circuitry. In addition, silicon photodiodesare physically small, which enables them to be readily incorporated intoa system for use with a membrane-based device. If silicon photodiodesare used, then the wavelength range of the emitted signal may be withintheir range of sensitivity, which is 400 to 1100 nanometers. Anothersuitable detector is a CdS (cadmium sulfide) photoconductive cell, whichhas the advantage of having a spectral sensitivity similar to that ofhuman vision that may make rejection of the reflected emitted radiationeasier.

Referring again to FIG. 2, the detector 54 is positioned adjacent to thesupport 21 and the EL illumination source 52 is positioned adjacent tothe second surface 15 of the chromatographic medium 23 Likewise, thedetector 54 may be positioned adjacent to the second surface 15 of thechromatographic medium 23 and the EL illumination source 52 may bepositioned adjacent to the support 21. Thus, the EL illumination source52 may emit light simultaneously onto the detection and calibrationzones 31 and 32 and the detector 54 may likewise also simultaneouslyreceive a detection signal from the detection probes at the detectionand calibration zones 31 and 32. Alternatively, the EL illuminationsource 52 may be constructed to successively emit light onto thedetection zone 31 and the calibration zone 32. In addition, a separateillumination source and/or detector (not shown) may also be used for thecalibration zone 32. It should also be understood that the ELillumination source 52 and detector 54 may be positioned on the sameside of the assay device 20, such as both being adjacent to thechromatographic medium 23.

To improve the signal-to-noise ratio of the optical detection systemwithout the need for certain types of complex and expensive opticalcomponents, such as lenses or other light guiding elements, the distanceof the EL illumination source 52 and/or detector 54 from the assaydevice 20 may be minimized. For instance, as shown in FIG. 3 a, light(indicated by directional arrows) traveling a relatively large distancetends to diffuse, thereby causing some photons to miss the test sampleor the detector 54 To reduce light scattering, lenses may be employed tofocus the light in the desired direction, such as shown in FIG. 3 b.However, as shown in FIGS. 3 c and 3 d, the need for such expensive andcomplex equipment may be reduced by simply moving the EL illuminationsource 52 and/or detector 54 closer to the assay device 20. The use of ashorter light path results in less diffusion of the light. For example,FIG. 3 c illustrates an embodiment in which the EL illumination source52 is positioned closer to the assay device 20, and FIG. 3 d illustratesan embodiment in which both the EL illumination source 52 and detector54 are positioned closer to the assay device 20. Thus, in someembodiments, the EL illumination source and/or detector 54 may bepositioned less than about 5 millimeters, in some embodiments less thanabout 3 millimeters, and in some embodiments, less than about 2millimeters from the assay device 20. For example, the EL illuminationsource 52 may be laminated directly to the support 21. Likewise, as willbe discussed in more detail below, the EL illumination source 52 and/ordetector 54 may, in some cases, directly contact the chromatographicmedium 23 For example, the EL illumination source 52 may carry themedium 23, thereby also functioning as its support. In other cases,however, it may be desired to keep the EL illumination source 52 and/ordetector 54 at a distance that is large enough to avoid contamination ofany biological reagents. For example, the EL illumination source 52and/or detector 54 may sometimes be positioned at a distance of fromabout 1 to about 3 millimeters from the assay device 20.

In FIG. 2, the EL illumination source 52 is shown as a component that isseparate from the assay device 20. However, the present invention alsocontemplates embodiments in which the illumination source is integralwith the assay device 20. For example, in some embodiments, the support21 may function simultaneously as a physical carrier for thechromatographic medium 23 and also as the EL illumination source for theoptical detection system. The use of an EL device as the support 21provides a substantial benefit to the resulting optical detection systemby eliminating the need for additional light sources, which are oftencostly and lead to overly complex and space-consuming systems. That is,the EL device may be laminated to the chromatographic medium 23 andsimultaneously function as the support 21 and light source for theoptical detection system. The EL device may be selected to possess acertain degree of flexibility that allows it to be readily manipulatedand/or cut into the desired shape and size for the assay device 20. Onecommercially available EL device that has enough strength andflexibility for use as the support 21 is a lamp kit available fromGraphic Solutions Int'l, LLC of Burr Ridge, Ill. under the name“Proto-Kut.”

One particular embodiment of the present invention in which an EL deviceis employed as the support for the assay device is shown in FIG. 4.Specifically, an assay device 220 is depicted that includes achromatographic medium 223, an EL device 221, an absorbent pad 228, anda conjugate pad 222. The medium 223 has a first surface 212 and a secondsurface 214, wherein the first surface 212 is positioned adjacent to theEL device 221. A detection zone 231 and calibration zone 232 are definedby the medium 223 for providing detection and calibration signals.Further, a detector 254 positioned adjacent to the second surface 214 ofthe medium 223. In this particular embodiment, the EL device 221functions as both the illumination source and the support for the medium223. Leads 256 for the EL device 221 are connected to a driver circuit260 via wiring, which in turn, is connected to a power source 266. Thedetails of the driver circuit 260 and power source 266 depend on therequirements of the particular EL device. For example, because the ELdevice 221 may be relatively small due to the corresponding small sizeof the assay device 220, a low voltage circuit and battery power sourcemay be employed to reduce the cost and complexity of the system.However, higher voltage circuits may also be used, such as a drivercircuit that converts DC voltage into an AC output for driving the ELdevice 221. Such AC inverters may generate around 60 to 300 volts AC at50 to 5000 Hertz.

As mentioned above, the EL illumination device of the present inventionmay provide diffuse illumination. In this manner, the reliance oncertain external optical components, such as diffusers, may be virtuallyeliminated. Regardless, such optical components may nevertheless beutilized in some embodiments of the present invention. If utilized,separate optical components may be used for the EL illumination source52 and detector 54, or they may share common optical components. Forexample, optical diffusers may be utilized in the present invention toscatter light in a certain direction, such as toward and/or away fromthe detection zone. Suitable optical diffusers may include diffusersthat scatter light in various directions, such as ground glass, opalglass, opaque plastics, chemically etched plastics, machined plastics,and so forth. Opal glass diffusers contain a milky white “opal” coatingfor evenly diffusing light, thereby producing a near Lambertian source.Other suitable light-scattering diffusers include polymeric materials(e.g., polyesters, polycarbonates, etc.) that contain a light-scatteringmaterial, such as titanium dioxide or barium sulfate particles. In otherembodiments, holographic diffusers may be utilized that both homogenizeand impart predetermined directionality to light rays emanating from theillumination source. Such diffusers may contain a micro-sculpted surfacestructure that controls the direction in which light propagates.Examples of such holographic diffusers are described in more detail inU.S. Pat. No. 5,534,386 to Petersen, et al., which is incorporatedherein in its entirety by reference thereto for all purposes.

Optical filters (not shown) may also be disposed adjacent to the ELillumination source 52 and/or detector 54 The optical filters may havehigh transmissibility in a desired wavelength range(s) and lowtransmissibility in one or more undesirable wavelength band(s) to filterout undesirable wavelengths from the EL illumination source 52. Inluminescent detection systems, for instance, undesirable wavelengthranges may include those wavelengths that produce detectable sampleautofluoresence and/or are within about 25 to about 100 nanometers ofexcitation maxima wavelengths and thus are potential sources ofbackground noise from scattered excitation illumination. Severalexamples of optical filters that may be utilized in the presentinvention include, but are not limited to, dyed plastic resin or gelatinfilters, dichroic filters, thin multi-layer film interference filters,plastic or glass filters, epoxy or cured transparent resin filters. Inone embodiment, the detector 54 and/or EL illumination source 52 may beembedded or encapsulated within the filter.

In addition, a lens may also be used to collect and focus light. Oneparticular embodiment of the present invention utilizes a micro-lens tofocus light toward the test sample and/or detector 54. Suitablemicro-optic lenses include, but are not limited to, gradient index(GRIN) lenses, ball lenses, Fresnel lenses, and so forth. For example, agradient index lens is generally cylindrical, and has a refractive indexthat changes radially with a parabolic profile. A ball lens is generallyspherical, and has a refractive index that is radially constant. Becauseof their relatively small size, such micro-lenses may be particularlyadvantageous in the present invention. Any of a variety of well-knowntechniques may be utilized to form the micro-lens. For example,micro-lenses may be formed by submerging a substrate (e.g., silicon orquartz) into a solution of alkaline salt so that ions are exchangedbetween the substrate and the salt solution through a mask formed on thesubstrate, thereby obtaining a substrate having a distribution ofindexes of refraction corresponding to the pattern of the mask. Inaddition, a photosensitive monomer may be irradiated with ultravioletrays to polymerize an irradiated portion of the photosensitive monomer.Thus, the irradiated portion bulges into a lens configuration under anosmotic pressure occurring between the irradiated portion and thenon-irradiated portion. In another embodiment, a photosensitive resinmay be patterned into circles, and heated to temperatures above itssoftening point to enable the peripheral portion of each circularpattern to sag by surface tension. This process is referred to as a“heat sagging process.” Further, a lens substrate may simply bemechanically shaped into a lens. Still other suitable techniques forforming a micro-lens or other micro-optics are described in U.S. Pat.No. 5,225,935 to Watanabe, et al.; U.S. Pat. No. 5,910,940 to Guerra;and U.S. Pat. No. 6,411,439 to Nishikawa, which are incorporated hereinin their entirety by reference thereto for all purposes.

Further, a mask, such as a black coating or dye, may be utilized toprevent light from passing through one or more sections of the assaydevice 20. Light guiding elements may also be utilized to direct lightin a desired direction, such as a single optical fiber, fiber bundle,segment of a bifurcated fiber bundle, large diameter light pipe, planarwaveguide, attenuated total reflectance crystal, dichroic mirror, planemirror or other light guiding elements. Still other examples ofoptically functional materials that may be used in the present inventiondescribed in U.S. Pat. No. 5,827,748 to Golden; U.S. Pat. No. 6,084,683to Bruno, et al.; U.S. Pat. No. 6,235,241 to Catt, et al.; U.S. Pat. No.6,556,299 to Rushbrooke, et al.; and U.S. Pat. No. 6,566,508 to Bentsen,et al., which are incorporated herein in their entirety by referencethereto for all purposes.

If desired, the optical properties of the assay device itself may beselectively tailored to the optical requirements of the detectionsystem. For example, referring again to FIG. 2, one embodiment of thepresent invention employs selective control of the support 21 tooptimize the performance of the optical detection system. In oneparticular embodiment, for example, the support 21 is opticallytransmissive to allow for light to travel from the EL illuminationsource 52 to the detector 54 In addition, the support 21 may function asa diffuser for the EL illumination source 52 and/or detector 54 toimprove the signal-to-noise ratio of the optical detection system. Thesupport 21 may also function as an optical filter of the detectionsystem. Thus, in the illustrated embodiment, light from the ELillumination source 52 is absorbed by probes (not shown) present at thedetection zone 31 and/or calibration zone 32. The probes produce asignal that is attenuated by the optical filter before reaching thedetector 54. The optical filter may, for example, have hightransmissibility in the emission wavelength range(s) and lowtransmissibility in one or more undesirable wavelength band(s) to filterout undesirable wavelengths from the detector 54. The optical detectionsystem may also include an additional optical filter (not shown)positioned between the EL illumination source 52 and the chromatographicmedium 23. This additional optical filter may have high transmissibilityin the excitation wavelength range(s) and low transmissibility in one ormore undesirable wavelength band(s). Alternatively, an additionaloptical filter may be integrated into the EL illumination source 52and/or detector 54. The support 21 may also posses other desirableoptical qualities. For example, the support 21 may contain a mask, lightguiding element, lens, etc. In some cases, when employed in the support21, it is desired that “micro-optic” elements are utilized. Micro-opticelements generally have a size less than about 2 millimeters and arearranged in one or two dimensions. Due to their small size, micro-opticelements may be more readily utilized in the support 21.

When the support 21 is optimized for a particular optical property, thematerial(s) used for forming the support 21 may be selected to possessthe desired optical property. Alternatively, the desired opticallyfunctional material may simply be applied to the support 21 beforeand/or after forming the assay device 20. Such an optically functionalmaterial may be applied to the support 21 in a variety of ways. Forexample, the optically functional material may simply be dyed or coatedonto one or more surfaces of the support 21. When applied in thismanner, the optically functional material may cover only a portion or anentire surface of the support 21. In one embodiment, for example, theoptically functional material is applied to a portion of the support 21that corresponds to the detection zone 31 and/or calibration zone 32. Inthis manner, the optically functional material may enhance the detectionor calibration signals produced by the assay device 20 during use.Alternatively, the optically functional material may also beincorporated into the structure of the support 21. For example, internaloptics may be formed using known techniques, such as embossing,stamping, molding, etc.

In accordance with certain embodiments of the present invention, theoptical detection system may also employ various other components thatenhance the detection sensitivity of the analyte. For example, thedetection system may sometimes employ a sample holder for the assaydevice. Referring to FIGS. 5-8, for example, various embodiments of anoptical detection system that employs such a sample holder will now bedescribed in more detail. FIG. 5, for instance, illustrates oneembodiment of a sample holder 400 that may be employed in the opticaldetection system of the present invention. As shown, the sample holder400 includes a lower portion 402 and an upper portion 403. The upperportion 403 is capable of movement about a hinge 404 so that it may bepositioned in an open position (FIGS. 5A and 5B) and a closed position(FIG. 5C). Further, the sample holder 400 may include an upper latch 413that mates with a lower latch 415 for securing the holder 400 in itsclosed position. A handle 417 may also be provided to allow a user tomore readily grip the holder 400.

As shown, one or more assay strips 405 may be disposed within aninterior of the sample holder 400 defined between the lower portion 402and upper portion 403. In this particular embodiment, the support card(not shown) of the assay strips 405 is also laminated to EL devices 412.This allows the EL devices 412 to be positioned close to the assaystrips 405 during use to optimize the signal-to-noise ratio of theoptical detection system. The EL devices 412 may be placed intoelectrical contact with leads in a variety of different ways. Forexample, the lower surface of the EL devices (e.g., cathode-side) may beplaced adjacent to eight (8) holes 409, although any number of holes mayof course be utilized. Referring to FIGS. 6 and 7, these holes 409 maybe positioned adjacent to eight (8) corresponding leads 313 (only 3 ofwhich are shown in FIGS. 6 and 7) of a cartridge 300. Specifically, auser may align one end 419 of the holder 400 with a sample port 315defined by a body portion 310 of the cartridge 300, and thereafter slidethe sample holder 400 through the sample port 315 via parallel tracks319 until the holes 409 are positioned over the leads 313. In thismanner, the lower side (e.g., cathode-side) of the EL devices 412 isplaced into electrical contact with the leads 313. Although notspecifically illustrated, an upper surface of the EL devices 412 (e.g.,anode-side) may also extend beyond the assay strips 405 and placed intoelectrical contact with leads. For example, leads (not shown) may bedisposed on the inner surface of the upper portion 403 of the sampleholder 400 (FIG. 5) so that when the holder is closed, the leads contactthe extended portion of the upper surface of the EL devices 412. Thus,during use, the EL devices 412 generate illumination that contactsdetection probes located on the assay strips 405. The detection probesproduce a detection signal that travels through an upper window 406 ofthe sample holder 400 and an upper window 326 of the cartridge 300before reaching a lens 501 of a camera 500.

If desired, as shown in FIG. 8, the above-referenced components may becontained within an enclosure 600 that is not transmissive to theelectromagnetic radiation emitted by the EL device or registered by thedetector to optically isolate the system. In the illustrated embodiment,for example, the sample holder 400 (FIG. 5), the cartridge 300 (FIG. 6),and the camera 500 (FIG. 7) are positioned within the enclosure 600.Although shown as having an oval shape, it should be understood that anyother suitable shape and/or size may be employed, such as circular,square, rectangular, etc. Further, as would be readily recognized bythose skilled in the art, other optical components may also be utilizedand optionally contained within the enclosure 600, such as electroniccircuitry, microprocessors, displays, mirrors, optical filters, lenses,and so forth.

Regardless of the specific manner in which the optical detection systemis formed, qualitative, quantitative, or semi-quantitative determinationof the presence or concentration of an analyte may be achieved inaccordance with the present invention. For example, in one embodiment,the amount of the analyte may be quantitatively or semi-quantitativelydetermined by correlating the intensity of the signal, I_(s), of theprobes captured at the detection zone 31 with a predetermined analyteconcentration. In some embodiments, the intensity of the signal, I_(s),may also be compared with the intensity of the signal, I_(c), of theprobes captured at the calibration zone 32. The intensity of the signal,I_(s), may be compared to the intensity of the signal, I_(c). In thisembodiment, the total amount of the probes at the calibration zone 32 ispredetermined and known and thus may be used for calibration purposes.For example, in some embodiments (e.g., sandwich assays), the amount ofanalyte is directly proportional to the ratio of I_(s) to I_(c). Inother embodiments (e.g., competitive assays), the amount of analyte isinversely proportional to the ratio of I_(s) to I_(c). Based upon theintensity range in which the detection zone 31 falls, the generalconcentration range for the analyte may be determined.

A microprocessor may optionally be employed to convert the measurementfrom the detector 54 to a result that quantitatively orsemi-quantitatively indicates the presence or concentration of theanalyte. The microprocessor may include memory capability to allow theuser to recall the last several results. Those skilled in the art willappreciate that any suitable computer-readable memory devices, such asRAM, ROM, EPROM, EEPROM, flash memory cards, digital video disks,Bernoulli cartridges, and so forth, may be used in the presentinvention. Optical density (grayscale) standards may also be used tofacilitate a quantitative result as is well known in the art. Further,any known software may optionally be employed for data collection. Forexample, Logitech camera software may be used to collect data obtainedfrom a Logitech camera-based detector. After the images are saved, theymay be analyzed using any known commercial software package, such asImageQuant from Molecular Dynamics of Sunnyvale, Calif. If desired, theresults may be conveyed to a user using a liquid crystal (LCD) or LEDdisplay.

The present invention may be better understood with reference to thefollowing examples.

EXAMPLE 1

The ability to form an optical detection system with anelectroluminescent illumination source was demonstrated. Initially, anitrocellulose membrane (SHF-120, Millipore Corp. of Bedford, Mass.) wasprovided that was laminated to a Mylar® film support. The Mylar® filmwas attached directly to an electroluminescent (EL) device using atransparent adhesive obtained from Adhesives Research of Glen Rock, Pa.under the name “ARclear 8154.” Care was taken to ensure the absence ofbubbles, dust, and contaminants. The EL device was made by BKL, Inc. ofBurr Ridge, Ill., and had a size of 60 millimeters×300 millimeters. Inaddition, the EL device also had a dual, broad emission maxima of 482and 580 nanometers to give “white” light emission.

Goldline™ (a polylysine solution obtained from British BiocellInternational) was striped onto the membrane to form a calibration zone.Monoclonal antibody reactive toward C-reactive protein (BiosPacific,Inc., concentration of 1 milligram per milliliter) was immobilized onthe porous membrane to form a detection zone. The card was then driedfor 1 hour at a temperature of 37.5° C. After the card was removed fromthe oven, a cellulosic wicking pad (Millipore Co.) was attached to theend of the membrane closer to the calibration zone. The other end of thecard, typically used to attach conjugation and sample pads, was removed.The card was then sliced into strips (4 mm×60 mm in size). Carboxylatedblue latex beads (0.3 millimeters, Bang's Laboratories) were conjugatedto monoclonal antibody reactive toward C-reactive protein (BiosPacific,Inc., concentration of 1 milligram per milliliter). The conjugate wasmixed with various concentrations of C-reactive protein (CRP) serumstandard (Kamiya), put into a micro-well plate, and tested against thehalf sticks. The blue detection and control lines developed within aminute.

After drying at ambient conditions for 1 hour, the lateral flow stripswere loaded, four at a time, into a sample holder as shown in FIG. 5.The holder, when closed, immobilized the strips in such a manner thatexposed electrodes on the underside of the EL device were aligned withholes in the holder. As shown in FIGS. 7 and 8, the sample holder wasthen inserted into an enclosure that housed a camera system. The purposeof the enclosure was to optically isolate the system from the externalenvironment and ensure proper alignment between the camera and the assaydevices. The camera was a Logitech QuickCam 3000 obtained from LogitechInc. of Fremont, Calif., and used a USB connection to a standard desktopPC. The camera was fitted with a Finite Conjugate Micro Video ImagingLens (8 mm focal length, NT54-853) from Edmund Industrial Optics ofBarrington, N.J., and was positioned so that the surface of thenitrocellulose membrane was 47 millimeters from the imaging plane of theCCD. The EL device was powered by an AC power supply (ACM-500 fromBehlman, Hauppauge, N.Y.) at 100 V and 400 Hz. Spring-loaded contacts(70AD/Male/4-up, Bourns, Riverside, Calif.) were mounted inside of theenclosure and made electrical contact through holes in the sampleholder.

The images of the illuminated assay devices were collected and analyzedusing Visual Basic (VB) software. The VB software used an ActiveX module(QCSDK1) to control the Logitech Twain device driver, which in turncontrolled the CCD camera. A variety of image acquisition parameterswere controlled, including Brightness, Exposure, Gain, Saturation andWhite Balance. The values of each were: Brightness=204 (out of 256),Exposure= 1/300 sec, Gain=0 (out of 256), Saturation=120 (out of 256)and White Balance=120 (out of 256). Additionally, ten images were takenin succession and averaged to reduce noise. After the average image wasacquired, regions of interest (ROI) for analysis (i.e., the bands andtheir surroundings) were identified by placing and sizing rectanglesaround the features and representative background areas. The averagevalue of the pixels in the background region was calculated and used tonormalize the pixels in the ROI. The data was also corrected withcalibration data derived from images of blank strips. The averageintensity of the pixels within the region of interest and the area ofthe pixels were calculated using the trapezium method. The results weredisplayed on the screen and charts were drawn with the intensity data.FIG. 9 illustrates the response curve obtained from variousconcentrations of C-reactive protein.

EXAMPLE 2

The ability to form an optical detection system with anelectroluminescent illumination source was demonstrated. Initially, anitrocellulose membrane (SHF-120, Millipore Corp. of Bedford, Mass.) wasprovided that was laminated to a Mylar® film support. The Mylar® filmwas attached directly to an electroluminescent (EL) device using atransparent adhesive obtained from Adhesives Research of Glen Rock, Pa.under the name “ARclear 8154.” Care was taken to ensure the absence ofbubbles, dust, and contaminants. The EL device was made by BKL, Inc. ofBurr Ridge, Ill., and had a size of 60 millimeters×300 millimeters. Inaddition, the EL device also had an emission maxima of 525 nanometers togive “green” light emission.

Monoclonal antibody reactive toward C-reactive protein (BiosPacific,Inc., concentration of 1 milligram per milliliter) was conjugated tocolloidal gold particles having a size of 40 nanometers. The conjugatewas then diluted in 2-millimolar hydrated sodium borate (Borax, pH 7.2)and 50% sucrose (final 10% sucrose). The conjugate was sprayed onto5-millimeter wide glass fiber strips (Millipore GF33) at a rate of 5microliters per centimeter and at a bed speed of 5 centimeters persecond using a Kinematic 1600 dispenser. The sprayed conjugate stripswere allowed to dry overnight at less than 20% relative humidity and atroom temperature. The conjugate strips were then heat-sealed intoimpervious bags with desiccant. Goat-Anti-Mouse Antibody (GAM) wasdiluted in phosphate-buffered saline (PBS) to 0.1 milligram permilliliter and striped onto nitrocellulose membranes (HF120, Millipore)using the Kinematic 1600 dispenser at a dispense rate of 1 microlitersper centimeter and at a bed speed of 5 centimeters per second.Biogenesis CRP (KC202004A, 2.59 milligrams per milliliter) was alsostriped neat below the GAM test line. The cards were left to dry at 37°C for 1 hour. Upper wick and conjugate bands were attached with a3-millimeter overlap and striped onto the nitrocellulose membrane. CRPstandard (Scipac) was diluted in PBS to give the following final CRPconcentrations: 100, 20, and 0 micrograms per milliliter. Two hundredmicroliters of each standard solution was applied to a strip. Afterdrying at ambient conditions for 1 hour, several of the lateral flowstrips were analyzed as described in Example 1. The results are setforth below in Table 1. TABLE 1 Results of CRP Analysis SampleConcentration (μg/ml) Peak Area Peak Intensity 100 0.52 0.60 20 1.800.66 0 6.02 0.99

EXAMPLE 3

The ability to form an optical detection system with anelectroluminescent illumination source was demonstrated. The EL devicewas made by BKL, Inc. of Burr Ridge, Ill., and had a size of 60millimeters×300 millimeters. In addition, the EL device also had anemission maxima of 525 nanometers to give “green” light emission. The ELdevice was powered by an AC power supply (ACM-500 from Behlman,Hauppauge, N.Y.) at 100 V and 400 Hz. The EL device was cut into 4 mm×60mm strips that were inserted into an enclosure as shown in FIGS. 5-8using spring-loaded contacts (70AD/Male/4-up, Bourns, Riverside, Calif.)to make electrical contact through holes in the sample holder. Theenclosure housed two blue-enhanced silicon photodiodes (PDB-V601,Photonic Detectors of Simi Valley, Calif.). The purpose of the enclosurewas to optically isolate the system from the external environment andensure proper alignment between the photodiodes and assay devices. Thephotodiodes were positioned so that they would be 100 micrometers fromthe surface of the membrane upon insertion. The leads from the twophotodiodes were connected such that the diodes were wired in series,but reversed with respect to one another (i.e., the cathodes of the twophotodiodes were soldered together). The two anode wires were connectedto the probe leads of a multimeter (123 Industrial Scopemeter, Fluke,Everett, Wash.).

While the invention has been described in detail with respect to thespecific embodiments thereof, it will be appreciated that those skilledin the art, upon attaining an understanding of the foregoing, mayreadily conceive of alterations to, variations of, and equivalents tothese embodiments. Accordingly, the scope of the present inventionshould be assessed as that of the appended claims and any equivalentsthereto.

1. An optical detection system for detecting the presence or quantity ofan analyte within a test sample, said system comprising anelectroluminescent illumination source that provides electromagneticradiation, said electromagnetic radiation being capable of causing theproduction of a detection signal that correlates to the presence orquantity of the analyte.
 2. The optical detection system of claim 1,wherein said electroluminescent illumination source is a dispersion orthin-film device.
 3. The optical detection system of claim 1, whereinsaid electroluminescent illumination source contains a luminescent layersandwiched between at least two electrodes.
 4. The optical detectionsystem of claim 3, wherein at least one of said electrodes containscarbon.
 5. The optical detection system of claim 3, wherein at least oneof said electrodes contains a metal selected from the group consistingof aluminum, gold, silver, copper, platinum, palladium, iridium, andalloys thereof.
 6. The optical detection system of claim 3, wherein atleast one of said electrodes contains an inorganic oxide.
 7. The opticaldetection system of claim 6, wherein said inorganic oxide is selectedfrom the group consisting of indium oxide, indium tin oxide, tin oxide,antimony tin oxide, and combinations thereof.
 8. The optical detectionsystem of claim 7, further comprising a dielectric layer positionedbetween said luminescent layer and at least one of said electrodes. 9.The optical detection system of claim 3, wherein said luminescent layercontains phosphor particles.
 10. The optical detection system of claim1, further comprising: an assay device that includes a chromatographicmedium in communication with detection probes, said detection probesbeing capable of producing said detection signal; and a detector capableof registering said detection signal produced by said detection probes.11. The optical detection system of claim 10, wherein saidelectroluminescent illumination source and said detector are positionedon the same side of said assay device.
 12. The optical detection systemof claim 10, wherein said electroluminescent illumination source andsaid detector are positioned on opposing sides of said assay device sothat said chromatographic medium is positioned in the electromagneticradiation path defined between said electroluminescent illuminationsource and said detector.
 13. The optical detection system of claim 10,wherein said chromatographic medium is a porous membrane.
 14. Theoptical detection system of claim 10, wherein said chromatographicmedium includes a fluidic channel.
 15. The optical detection system ofclaim 10, wherein a receptive material is immobilized within a detectionzone defined by said chromatographic medium, said receptive materialbeing configured to bind to at least a portion of said detection probesor complexes thereof.
 16. The optical detection system of claim 10,wherein said chromatographic medium is carried by saidelectroluminescent illumination source.
 17. The optical detection systemof claim 16, wherein said electroluminescent illumination source islaminated to a support for said chromatographic medium.
 18. The opticaldetection system of claim 16, wherein said electroluminescentillumination source is laminated to said chromatographic medium.
 19. Theoptical detection system of claim 16, wherein an optically transparentadhesive is used to laminate said electroluminescent illumination sourceto said chromatographic medium, to a support for said chromatographicmedium, or combinations thereof.
 20. The optical detection system ofclaim 10, wherein said assay device is contained within a sample holder.21. The optical detection system of claim 20, wherein saidelectroluminescent illumination source is also positioned within saidsample holder.
 22. The optical detection system of claim 21, furthercomprising a cartridge that contains leads for said electroluminescentillumination source, said cartridge defining a port that receives saidsample holder.
 23. An optical detection system for detecting thepresence or quantity of an analyte within a test sample, said systemcomprising: an assay device that includes a porous membrane incommunication with detection probes, said detection probes being capableof producing a detection signal; an electroluminescent illuminationsource capable of providing electromagnetic radiation that causes saiddetection probes to produce said detection signal; and a detectorcapable of registering said detection signal produced by said detectionprobes.
 24. The optical detection system of claim 23, wherein saidelectroluminescent illumination source is a dispersion or thin-filmdevice.
 25. The optical detection system of claim 23, wherein saidelectroluminescent illumination source contains a luminescent layersandwiched between at least two electrodes.
 26. The optical detectionsystem of claim 23, further comprising a dielectric layer positionedbetween said luminescent layer and at least one of said electrodes. 27.The optical detection system of claim 23, wherein saidelectroluminescent illumination source and said detector are positionedon the same side of said assay device.
 28. The optical detection systemof claim 23, wherein said electroluminescent illumination source andsaid detector are positioned on opposing sides of said assay device sothat said chromatographic medium is positioned in the electromagneticradiation path defined between said electroluminescent illuminationsource and said detector.
 29. The optical detection system of claim 23,wherein said porous membrane is carried by said electroluminescentillumination source.